Editing 293FT Culture and Protein Production (section)

=== TROUBLESHOOTING<br> ===

Problem: The cells do not grow well. <br>[Step 9] <br>Solution: If cells do not grow with the expected doubling time, consider the following: <br>1. The cultureware is not sufficiently clean. Glassware for all culture manipulations and reagent preparations must be rinsed at least three times with high-purity H2O. <br>2. The H2O used may not be sufficiently pure. Check the source of the ultrapure H2O. <br>3. Check that the filtration system (if applicable) used to make the ultrapure H2O is functioning properly. <br>4. Cell aggregation can give a false impression of slower growth rates. Check regularly that the orbital shaker is at the right speed and that the incubator is at the correct temperature and is supplying the prescribed amount of CO2. <br>5. The cells may be too old. The number of cell passages indicates the age of the cells in service. A new cryovial can be thawed every 3 mo, on average. <br>6. Verify expiration dates, culture medium lots, and batches of additives used (e.g., Pluronic F-68 and antibiotics). <br>7. Check for mycoplasma contamination; if positive, thaw new cells<br>

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